This project's long-term goal is to use a well-characterized model system to define processes governing tolerance versus autoimmunity to self-antigens. To this end, we have developed lineages of transgenic mice expressing the influenza virus hemagglutinin (HA) as a nominal self-antigen, and have been analyzing the extent and basis by which they induce CD4+ T and B cell tolerance to the HA. In the most recent funding period, we showed that mice expressing HA driven by a MHC Class II promoter (HACII mice) and co- expressing HA-specific CD4+ T cell receptors (TCRxHACII mice) spontaneously develop autoimmunity, with inflammatory arthritis as the prominent disease manifestation. We showed that arthritis development is driven by CD4+ T cells and depends on the synthesis of the neo-self HA molecule by antigen presenting cells (APCs). Moreover, the penetrance of arthritis could be modulated by varying the reactivity of CD4+ T cells for the HA, since the majority of mice in which the TCR recognizes HA as an agonist peptide (TS1xHACII mice) developed arthritis, whereas mice expressing a TCR with ~100-fold lower reactivity for the HA (TS1(SW)xHACII mice) developed arthritis with substantially lower penetrance. This proposal will use this model system to understand how interactions between autoreactive CD4+ T cells and a self-peptide synthesized by systemically distributed APCs can lead to the development of autoimmune arthritis. In Aim 1 we will analyze how reciprocal interactions with APCs synthesizing their target peptide promote arthritogenic CD4+ T cell development, and examine whether arthritis is associated with elevated target antigen expression and/or regional APC activation. We will also examine whether lymphopenia-induced CD4+ T cell proliferation contributes to arthritis development in TCRxHACII mice. In Aim 2 we will examine factors that can contribute to disease penetrance and severity in TCRxHACII mice. We will determine how CD4+ T cell frequency and/or B cell function can shape the severity and penetrance of arthritis, and examine whether the ability of cytokine blockade to modulate arthritis severity is sensitive to variations in the autoreactive CD4+ T cell repertoire. In Aim 3 we will determine whether arthritis development can be predicted or promoted in a low penetrance setting. We will examine whether differences in the CD4+ T cell repertoire or serum cytokine composition can prospectively identify individuals that will develop arthritis, and analyze how infections might increase the penetrance of arthritis among genetically susceptible individuals. These studies will provide fundamental insights into the mechanisms of immune repertoire formation and tolerance, will have general applicability to the processes of autoimmunity, and will use experimental models with direct relevance to the diagnosis and treatment of human rheumatoid arthritis. PUBLIC HEALTH RELEVANCE Statement Autoimmune diseases are a result of the immune system attacking the body's own cells and tissues, but how these diseases arise remains poorly understood. This proposal uses genetically-modified mice to analyze mechanisms and cellular processes than can lead to the development of autoimmune arthritis. These studies aim to develop novel insights into the autoimmune disease process that will facilitate the diagnosis and treatment of human autoimmune diseases, particularly rheumatoid arthritis.